H and L subunit chains of type A botulinum toxin are being purified separately. The purified preparations will be used as immunizing antigens to obtain anti-H and anti-L sera. These sera will be compared for relative efficiencies in protecting mice against challenge with intact type A toxin. Attempts will be made to determine which is the subunit by which toxin binds to nerve endings. By elimination, the other subunit can be inferred to be the pharmacologically active part of the toxin molecule. If sufficient binding subunit is given to a mouse, the animal's receptors for toxin could be saturated and the animal made resistant to challenges of all botulinum toxin types. The conversion of the unnicked to dichain molecular form is identified tentatively as resulting from cleavage of an arginyl peptide bond. Confirmation of the affected bond will be made and the role played by arginyl residues in toxicity will be studied. The two major fragments that are formed first in the controlled digestion of nicked toxin with trypsin will be isolated and tested to see if either has the part by which intact toxin attaches to nerve endings. As time permits, further attempts will be made to identify and isolate the enzymes of proteolytic botulinum cultures which change the synthesized, unnicked toxin molecule into dichains that predominate in purified toxin preparations.